Device for spraying a reagent for fast microbiological analysis

ABSTRACT

Device for spraying a reagent onto a support adapted to retain microorganisms on a predetermined surface. The device includes a spraying bell as well as a nozzle for emitting a jet of droplets of the reagent into a spraying chamber comprised by the bell, the device also including an absorbent pad mounted against the bell transversely to the jet and closing the chamber from the opposite side to the nozzle with the exception of a circular central opening provided in the pad, the diameter of the central opening being adapted to enable a portion of the jet, when the device faces the support and is at a predetermined distance therefrom, to pass through the central opening over its entire area and be deposited on the whole of the predetermined surface of the support.

BACKGROUND

The present invention concerns a spraying device for fastmicrobiological analysis.

At present, checking the microbiological quality of liquids, gases orsurfaces in the context of industrial and medical activities has toconform to strict standards.

On account of this, the industrial players and health authorities musthave tools at their disposal making it possible to detectmicrobiological contaminations as soon as possible, in order to be ableto correct these in good time and at reduced cost.

In practice, microbiological monitoring is carried out on a gel growthmedium where microorganisms, after having been collected on amicroporous membrane, are cultured until they have been rendered visibleto the naked eye.

The incubation periods vary from one microorganism to another but are ingeneral at least 24 hours and sometimes more for slower growthmicroorganisms (such as mycobacteria) or because the microorganisms havebeen stressed by environmental conditions.

To render the detection more rapid, another approach consists ofreducing the minimum duration of culture (or even, for somemicroorganisms, of eliminating it completely) by basing the detection ofthe microorganisms on their metabolic activity.

A universal metabolic marker, most commonly adenosine triphosphate (ATP)contained in living microorganisms, is measured by bringing it intocontact with a reagent revealing the presence of ATP by luminescence(termed a “bio luminescence reagent”) which enables the presence ofmicroorganisms to be noticed without having to wait for colonies to formon a gel growth medium and to become visible to the naked eye.

The quantity of light emitted is a function of the mass of ATP and thusthe number of microorganisms.

A device for detection by fast microbiology is already known which iscommercialized by the applicant under the name Milliflex Rapid®, andwhich comprises:

-   -   a station for filtering a volume of liquid onto a membrane so as        to capture the microorganisms that may be contained in the        liquid on the membrane;    -   a station for spraying a reagent revealing the presence of ATP        by luminescence facing which the operator places the membrane,        after the filtering step and after having rendered the ATP of        the microorganisms accessible (by a step of lysis of the        microorganisms for example), for the reagent to be deposited;        and    -   a station for measuring the quantity of light emitted in        response to the depositing of the reagent revealing the presence        of ATP by luminescence, facing which the operator places the        membrane, after the spraying step, the light emitted by the        membrane being collected by a CCD camera and processed and then        analyzed to detect the presence of microorganisms on that        membrane.

The spraying station is provided with a spraying device comprising asprayer spraying droplets emitted in the form of a jet onto the membranein the ambient air situated above the membrane.

SUMMARY

The invention concerns the provision of a device of the same type thatboth has better performance and is more practical while remainingreliable in terms of the risks of contaminations of the support toanalyze.

To that end it provides a device for spraying a reagent onto a supportfor the fast microbiological analysis of said support, said supportbeing adapted to retain microorganisms on a predetermined surface,characterized in that said device comprises a spraying bell as well as anozzle for emitting a jet of droplets of said reagent into a sprayingchamber comprised by said bell, said device also comprising an absorbentpad mounted against said bell transversely to said jet and closing saidchamber from the opposite side to said nozzle with the exception of acircular central opening provided in said pad, the diameter of saidcentral opening being adapted to enable a portion of said jet, when saiddevice faces said support and is at a predetermined distance therefrom,to pass through said central opening over its entire area and bedeposited on the whole of said predetermined surface of said support.

The spraying device according to the invention, provided with a bell andan absorbent pad used in combination, makes it possible to keep the(relatively volatile) droplets of the sprayed jet confined such that itis thus possible to group together the measuring station and thespraying station into the same analysis chamber.

More particularly, in the absence of effective confinement of the jet ofdroplets, such grouping together would be rendered impossible sincethose (relatively volatile) droplets would risk being deposited on theentire surface of the analysis chamber including the measuring station.As this chamber may be contaminated by extraneous ATP (ATP which isnaturally present on the surfaces), putting the reagent in contact withthat extraneous ATP would generate light which would risk perturbing theanalysis of that emitted by the support.

Furthermore, the pad of this device is shaped so as to collect thedroplets situated at the periphery of the jet and liable to enter intocontact with the bell. This is because it is particularly important totrap those droplets since, having entered into contact with the bell(which may also be contaminated by extraneous ATP), they constitutepotential sources of contamination of the support by extraneous ATP ifthe paths of those droplets were to end their travel on that support.

The pad thus makes it possible to select only the portion of the jet ofdroplets adapted to uniformly cover the whole of the surface of thesupport to analyze, without risk of contaminating it.

Furthermore, the diameter of the opening of the pad, which is a functionof the distance to the support from the pad, means that the beam ofsprayed droplets leaving by the opening of the pad over the entire areaof that opening uniformly covers the entire surface to treat of thesupport and solely that surface in order to avoid any risk ofcontamination by the droplets coming into contact with the environmentclose to the support.

According to features that are preferred for reasons of simplicity andconvenience of use, said spraying chamber is delimited by a wallsubstantially transverse to said jet, facing said pad and at the centerof which is disposed said nozzle, by a side wall of which one edge joinsto the periphery of said substantially transverse wall, as well as bysaid pad disposed against said side wall;

According to still other features that are preferred for the samereasons as above, said side wall has at least one frusto-conical portionagainst which is disposed said pad and of which the taper is adapted toorientate the droplets which are at the periphery of said jet towardssaid pad.

That frusto-conical portion enables the droplets which do not havesufficient energy to rebound to flow along that portion and then to beabsorbed by the pad and, for those which have sufficient speed torebound, to orientate the rebounds towards the pad itself but not in thedirection of its central opening such that those droplets are alsotrapped by the pad.

According to still other preferred features:

-   -   said side wall comprises, between said transverse wall and said        frusto-conical portion against which is disposed said pad, an        intermediate portion also adapted to orientate the droplets        which are at the periphery of said jet towards said pad and of        lesser taper than that of said frusto-conical portion against        which is disposed said pad;    -   said transverse wall comprises a frusto-conical portion of taper        between that of said intermediate portion and that of said        frusto-conical portion against which is disposed said pad;        and/or    -   said device also comprises a ring for retaining said pad against        said bell.

According to still other preferred features, at the center of said ring,there is provided a circular opening of greater diameter than thediameter of the opening provided in said pad.

The difference in diameter means that a portion of the pad is visiblefrom the support such that in case the droplets strongly rebound on thesupport they land and are trapped on the pad and not on the ring (sincein that case they would risk falling back onto the support, carryingextraneous ATP).

According to still other preferred features:

-   -   said ring has snap-fitting means adapted to cooperate with        complementary snap-fitting means comprised by said bell;    -   said device also comprises a reservoir and a pump disposed        between said nozzle and said reservoir;    -   said reservoir is slidingly mounted relative to said bell, said        pump being adapted to make a said jet of droplets come out of        said nozzle on sliding of said reservoir in the direction of        said bell;    -   said reservoir comprises a body provided with an annular bearing        collar; and/or

said reservoir has snap-fitting means adapted to cooperate withcomplementary snap-fitting means comprised by said bell.

BRIEF DESCRIPTION OF THE DRAWINGS

The features and advantages of the invention will appear from thefollowing description, given by way of preferred but non-limitingexample, with reference to the accompanying drawings in which:

FIG. 1 is a perspective view of a spraying device according to theinvention;

FIG. 2 is a view similar to FIG. 1 but with part of the device cut away;

FIG. 3 is an elevation view in section taken along a median plane ofsymmetry of the device and of a filter cassette provided with amicroporous membrane to analyze disposed under the device using a robotarm;

FIG. 4 is a similar view to FIG. 3 but in which a liquid reservoir ofthe device is represented in the position it occupies after slidingtowards a spraying bell of the device to actuate the ejection of a jetof droplets;

FIG. 5 is an enlarged view of the filter cassette and of the end of thespraying device which faces the cassette, in which arrows show differentpossible paths for the droplets of sprayed reagent; and

FIG. 6 is a perspective view of the device into which a protective plughas been fitted, this assembly being placed in a packaging tray.

DETAILED DESCRIPTION

The spraying device 1 illustrated in FIGS. 1 and 2 comprises a reservoir2, a confinement bell 3, a spraying unit 4, a porous pad 5 and a fixingring 6 (FIG. 2).

The reservoir 2 is slidingly mounted relative to the bell 3 andcomprises a body 10 of plastics material as well as a cover 11.

The body 10 has a first cylindrical portion 12, a second cylindricalportion 13, an annular collar 14 and two legs 15.

The cylindrical portion 12 delimits a housing 17 for receiving thereagent which is obturated by the cover 11.

The cylindrical portion 13 is situated in line with the portion 12 onthe opposite side to the cover 11 and has an outer diameter less thanthat of the portion 12.

A screw thread 16 is formed in the neighborhood of the opposite end ofportion 13 to that connected to portion 12.

The collar 14 is transversely attached to the rest of the body at thejunction of portions 12 and 13.

The two legs 15 which are diametrically opposite each other project fromthe opposite side of the collar to the cover 11 Each leg has a tooth 18projecting from the opposite side to the other leg 15.

The cover 11 has two ducts 20 and 21.

A filter unit 22 (and respectively 23) is screwed into duct 20 (andrespectively duct 21).

The filter unit 22 is an air filter whereas unit 23 is a liquid filter.

Means 24 for identification of the reagent and/or of the device may bestuck or engraved around the reservoir.

The confinement bell 3 is of plastics material formed as a single piece.

The bell comprises a first portion 31 (FIG. 3) delimiting a housing 32for receiving the spraying unit 4 and a second portion 33 delimitingtogether with the pad 5 a spraying and confinement chamber 34.

Portion 31 of the bell is formed from a first cylindrical portion 35,from a second cylindrical portion 36 and from an annular collar 37.

Portion 35, which is of greater diameter than portion 36, is joined byone end to that portion (portions 35 and 36 being in line with eachother) and by the other end to the collar 37, the latter being connectedtransversely to portion 35.

Collar 37 has two diametrically opposite oblong apertures 38 (FIG. 1)into which the legs 15 of the reservoir are snap fitted.

In the snap-fitted state of the reservoir in the bell, the collars 14and 37 face each other with the teeth 18 of the snap-fitting feet 15being situated on the opposite side of the collar 37 to the collar 14.

Portion 33 of the bell has a significantly flared first frusto-conicalportion 40, a second very slightly flared portion 41 (here this portionis practically cylindrical, its taper is practically nil) and a thirdportion 42 also frusto-conical and of moderate degree of flare, againstwhich the pad 5 is disposed.

The degree of taper of portion 40 is thus greater than that of portion41 and less than that of portion 42.

Cylindrical portion 35 is joined to portion 40 via portion 36.

Portion 40 forms a wall substantially transverse to the direction ofspraying of the reagent and faces the pad 5 whereas portions 41 and 42form a lateral wall extending from the periphery of portion 40 to thepad 5.

This portion also comprises an annular collar 43 transversely connectedto portion 41 as well as an annular lip 44 connected to the oppositeperipheral end of portion 42 to that joined to portion 41.

The bell is also provided with reinforcing fins 45 at portion 36 andreinforcing fins 46 at portion 42 that are attached to collar 43 (FIG.1).

The absorbent pad 5, visible in FIG. 2, is of cellulose wadding andtakes the form of a disc 50 in which a circular central opening 51 isformed. The pad is disposed against the lip 44 of portion 42 and closesthe spraying chamber 34 except for the central opening 51 which enablessaid chamber to communicate with the outside of the device.

The ring 6 is of plastics material and takes the form of a disc 60 inwhich a circular opening 61 is formed at its center and is provided witha snap-fitting collar 62 at its periphery.

The ring also has a series of cut-outs 63 at its periphery.

Pad 5 is disposed between the lip 44 and the ring 6 with the ring snapfitted against the lip.

The diameter of the opening 51 here is 40 mm whereas the diameter of theopening 61 is 65 mm such that the pad has a portion 52 (FIG. 2) hiddenby portion 60 and a portion 53 which projects inwardly of the bell andwhich is not covered by portion 60 of ring 6.

The spraying unit 4 comprises a pump 70 and a spraying nozzle 71 as wellas a screw-fitting plug 72 which are illustrated in FIG. 2.

Via a duct 73 the pump communicates on one side with the housing 17 ofthe reservoir and on the other side with the nozzle 71, the duct 73continuing through the nozzle so as to issue in the housing 34 of thebell (FIG. 2).

The plug 72 as well as the pump 70 are disposed in the housing 32 (FIG.3) within portion 35, with the plug 72 screwed into the screw thread 16of the cylindrical portion 13.

Nozzle 71 has a frusto-conical portion 77 housed in portion 36 of whichone end issues in the confinement housing 34 as well as an annularcollar 74 transversely connected to the opposite end of thefrusto-conical portion to that emerging in housing 34, which abutsagainst portion 36 (FIG. 3).

Nozzle 71 is provided to emit a jet of microscopic droplets bymechanical actuation of the pump 70 engaged by a sliding movement of thereservoir 2 towards the bell 3 while bearing on the collar 14 (forexample with the assistance of a robot arm).

Device 1 is delivered in the packaging illustrated in FIG. 6 and takesthe form of a rigid tray 75 provided with a cover (not shown), acylindrical plug 76 being engaged around the bell, the plug abuttingagainst the collar 43 in order to protect the confinement chamber.

A description will now be given with the help of FIGS. 3 to 5 of how thereagent contained in the reservoir of the device is deposited on asupport in the form of microscopic droplets.

The support illustrated here is a cassette 80 comprising a filtermembrane 81 having useful surface 82 corresponding to a diameter equalto 49 mm and a body 83 surrounding the membrane 81.

Once the filtration of the microorganisms has been made onto themembrane 81 and after having made the ATP of the microorganisms retainedon the surface 82 accessible (for example by a step of lysis of themicroorganisms), the cassette 80 is placed, with the assistance of arobot arm 85 (FIG. 3) under the spraying device 1 at a predetermineddistance (here 19 mm) from the absorbent pad 5. This device, from whichthe plug 76 has been removed beforehand, is fixedly held to a frame (notshown) of the analysis device.

A motorized unit (not shown) is then actuated to bear against the collar14 of the reservoir so as to make the reservoir 2 slide towards the bell3, the latter remaining immobile (FIG. 4), and so causing the actuationof the pump 70 to eject a jet of microscopic droplets into the chamber34 through the nozzle 71.

The motorized unit is then actuated to release the pressure it exerts onthe collar 14 so as to allow the reservoir 2 to resume its initialposition relative to the bell 3 (FIG. 3) so as to be ready to perform anew spraying operation.

The majority of the micro-droplets cross the bell through the openings51 and 61 without entering into contact with either the bell or theabsorbent pad, those droplets being deposited onto the support evenlyand homogenously over the entire useful surface 82 to be analyzed.

The diameter of the opening 51 and the distance from the pad 5 relativeto the surface 82 are thus determined on the basis of the spatialdimensions of the jet of droplets in order for a high proportion(approximately 90%) of those droplets to pass through the opening 51 ofthe pad 5 over the entire area of that opening before being deposited onthe entire surface 82.

A small proportion of the micro-droplets (the remaining 10%), situatedat the periphery of the jet, are stopped by the absorbent pad 5 byentering directly into contact with it or else after having encounteredthe internal surfaces of the portions 40 to 42 of the bell and afterhaving flowed along them to the pad 5 or after having rebounded on thatsurface so as to land on the pad 5.

More particularly and as illustrated in FIG. 5, in which differentpossible paths of the droplets are represented by the arrows A to H, thetaper of the portions 40 to 42 is chosen such that, whatever thelocation at which the droplet rebounds, it is directed after its reboundtowards portions 52 and 53 of the absorbent pad 5 and not in thedirection of the central opening 51.

This is because the coming into contact of the droplets with the bellleads to a risk of potential contamination by the extraneous ATP thatmay be present on the bell such that it is sought to trap in theabsorbent pad 5 all those droplets (paths A to C) in order to avoid themfalling back onto the membrane 81 carrying extraneous ATP which they mayhave captured on the bell.

It will be noted that the frusto-conical portion 40 is as flared aspossible in order to attempt to minimize the risks of contact with thedroplets.

Moreover portion 41 is practically cylindrical so as to reduce thedimensions of the bell.

The degree of taper of portion 42 and of portion 41 are chosen toorientate the rebounds of the droplets as desired without the bellassuming dimensions that are too large.

The major proportion of the droplets which pass through the opening 51is directly absorbed by the support (paths D and E) or rebounds slightlywithout coming back into contact with the device (path F).

However, where the rebound leads to a contact with the device (paths Gand H), it will be noted that the annular portion 53 of the absorbentpad projecting inwardly of the bell makes it possible in that case totrap those droplets which then come back into contact with the absorbentpad and not into contact with the ring 6 of which the opening 61 is ofgreater diameter than that of the opening 51 for that reason (since inthat case they then risk falling back onto the support, carryingextraneous ATP).

It can be noted that surface 82 and solely that surface receives thedroplets of the jet, path D being the most extreme path possible. Beyondthat, the droplets are captured by the pad such that no droplet riskscoming into contact with the body 83 of the cassette 80 and then fallingback onto the membrane 81, risking contaminating it with the extraneousATP.

The support thus treated is then available for the microbiologicalanalysis for example by moving the cassette 80 to the measuring station(not shown) within the same chamber in order to measure the quantity oflight emitted in response to putting the reagent in contact with the ATPof the microorganisms that may be present on the membrane using aphotomultiplier for example.

The device according to the invention may be used to treat severalmembranes, on each spraying operation the volume of liquid which wassprayed is replaced in the housing 17 by the air entering that housingthrough the pump 70.

It is also possible to regularly fill that housing by injecting a volumeof reagent through the filter unit 23 using a syringe, the unit 22forming a vent in that case.

Where the filter cassette is provided with a membrane of differentdiameter and thus of different useful surface, it is possible to adjustthe distance separating that membrane from the pad 5 by moving thedevice 1 vertically in order for the jet of droplets to cover the wholeof the surface of that membrane and solely that surface.

The device may be a single-use device but may also be used as many timesas necessary provided that the pad is not saturated with liquid.

As a variant the spraying device may also be used in any microbiologicalanalysis method requiring a pre-treatment step aimed at eliminating theextraneous ATP that the membrane contains by spraying that reagent onthe membrane before having rendered the ATP of the microorganismsaccessible (for example by lysis) in order for it to react and solelyeliminate the extraneous ATP, it being possible to carry out the lysisstep after neutralization of the sprayed reagent to eliminate theextraneous ATP.

In still another variant the spraying device may be used for any othertype of reagent intended for microbiological analysis which it is soughtto deposit uniformly over a surface while ensuring that the sprayedreagent is well confined.

The present invention is not limited to the embodiments described andrepresented but encompasses any variant form thereof.

The invention claimed is:
 1. A device for spraying a reagent onto asupport for the fast microbiological analysis of said support, saidsupport being adapted to retain microorganisms on a predeterminedsurface, said device comprising a spraying bell as well as a nozzle foremitting a jet of droplets of said reagent into a spraying chambercomprised by said bell, said device also comprising an absorbent padmounted against said bell transversely to said jet and closing saidchamber from the opposite side to said nozzle with the exception of acircular central opening provided in said pad, the diameter of saidcentral opening being adapted to enable a portion of said jet, when saiddevice faces said support and is at a predetermined distance therefrom,to pass through said central opening over its entire area and bedeposited on the whole of said predetermined surface of said support. 2.A device according to claim 1, wherein said spraying chamber isdelimited by a wall substantially transverse to said jet, facing saidpad and at the center of which is disposed said nozzle, and by a sidewall of which one edge joins to the periphery of said substantiallytransverse wall, as well as by said pad disposed against said side wall.3. A device according to claim 2, wherein said side wall has at leastone frusto-conical portion against which is disposed said pad and ofwhich the taper is adapted to orientate the droplets which are at theperiphery of said jet towards said pad.
 4. A device according to claim3, wherein said side wall comprises, between said transverse wall andsaid frusto-conical portion against which is disposed said pad, anintermediate portion also adapted to orientate the droplets which are atthe periphery of said jet towards said pad and of lesser taper than thatof said frusto-conical portion against which is disposed said pad.
 5. Adevice according to claim 4, wherein said transverse wall comprises afrusto-conical portion of taper between that of said intermediateportion and that of said frusto-conical portion against which isdisposed said pad.
 6. A device according to any one of claims 1 to 5,further comprising a ring for retaining said pad against said bell.
 7. Adevice according to claim 6, wherein, at the center of said ring, thereis provided a circular opening of greater diameter than the diameter ofthe opening provided in said pad.
 8. A device according to claim 6,wherein said bell has a portion having a lip, and wherein said ring hassnap-fitting means adapted to cooperate with said lip which comprisescomplementary snap-fitting means.
 9. A device according to claim 1,further comprising a reservoir and a pump disposed between said nozzleand said reservoir.
 10. A device according to claim 9, wherein saidreservoir is slidingly mounted relative to said bell, said pump beingadapted to make a said jet of droplets come out of said nozzle onsliding of said reservoir in the direction of said bell.
 11. A deviceaccording to claim 10, wherein said reservoir comprises a body providedwith an annular bearing collar.
 12. A device according to any one ofclaims 9 to 11, wherein said bell comprises a collar, and wherein saidreservoir has snap-fitting means adapted to cooperate with complementarysnap-fitting means on said collar.